A single HIV vaccine in human clinical trials demonstrated low level and short-term effectiveness, RV144. This vaccine used Pox virus prime - protein boost methodology, which because of the live virus prime, presents difficulties for large scale production and safety. We have developed a new approach to the prime portion of an HIV vaccine that uses nucleoside modified mRNA composed entirely of physiologic components that is simple and cost effective to produce, does not require a cold chain, and should have no adverse events due to the mRNA. The mRNA is complexed in a lipid nanoparticle (LNP) whose formulation has entered phase 3 clinical trials for siRNA with no adverse events due to the lipids and is delivered by the intradermal route. To complement this novel method of priming an immune response, we have also developed a new immunogen that is an HIV envelope that does not require CD4 binding to infect a cell (CD4 independent). This envelope has a more open conformation, and we have demonstrated that it induces higher levels of Env-specific IgG, Tier-1 and 2 neutralization, and V1/V2 directed responses compared to their parents. In 3 specific aims, we will developed a new CD4 independent immunogen from a transmitted/founder clade C virus, compare it to already produced CD4i immunogens in the rabbit model and downselect for study in Rhesus macaques. We will also analyze boost proteins, comparing gp120 monomers to SOSIP trimers, in rabbits for use as a boost in macaques. We will then compare single immunogen prime-boost to multi-genic vaccination. We believe this new vaccine approach will be effective, in that it will develop high levels of Env-specific IgG, Tier-1 and 2 neutralization, and V1/V2 directed responses. In addition, we have demonstrated that the nucleoside modified mRNA-LNP prime induces very high levels (half of the total antigen-specific CD4+ T cell response) of antigen-specific T follicular helper (Tfh) cell that are critical in both the generation of high avidity IgG and long term memory, both of which are critical elements in the development of an effective HIV vaccine. This grant will move a new approach to vaccine priming and optimize immunogens for the prime and boost for testing in rabbits and Rhesus macaques. The data generated will be capable of moving this vaccine approach to clinical trial development with a minority of additional experiments.